We mutagenized the X. tropicalis genome with Gamma Ray irradiation of sperm and subsequent in vitro fertilization. Sperm mutagenesis has a few advantages. First sperm can be treated with a much higher dose of mutagen than eggs, embryos, or adults. Sperm have the least amount of non-chromosomal material to be damaged. Gamma rays create chromosomal deletions either small or large or chromosomal rearrangements such as translocations or inversions. In addition sperm mutagenesis is easy to do.

One disadvantage of sperm mutagenesis is that the animals produced are mosaic. Therefore phenotypes from the F1 animals may not show simple mendelian genetics. On the other hand, these mosaic F1 animals may be able to harbor more mutations without lethal dominant effects.

In our hands, gamma ray exposure results in a stereotypical gastrulation defect that is dose dependent. We have tried a number of different doses although the optimal dose remains uncertain.

We recommend no cysteine or only a very light cysteine treatment just to separate the eggs. There will be a high mortality in this batch of embryos. It is considerably easier to simply place the embryos at low density and then just scoop out those embryos that hatch and are viable. Otherwise it would be hours of sorting dying embryos.